University of Wisconsin Milwaukee UWM Digital Commons Theses and Dissertations May 2017 Cyanobacteria and Cyanotoxin Ecology in Lakes and Drinking Water Chelsea Weirich University of Wisconsin-Milwaukee Follow this and additional works at: https://dc.uwm.edu/etd Part of the Environmental Health Commons Recommended Citation Weirich, Chelsea, "Cyanobacteria and Cyanotoxin Ecology in Lakes and Drinking Water" (2017). Theses and Dissertations. 1553. https://dc.uwm.edu/etd/1553 This Dissertation is brought to you for free and open access by UWM Digital Commons. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of UWM Digital Commons. For more information, please contact [email protected]. CYANOBACTERIA AND CYANOTOXIN ECOLOGY IN LAKES AND DRINKING WATER by Chelsea Weirich A Dissertation Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Environmental Health Sciences at The University of Wisconsin-Milwaukee May 2017 ABSTRACT CYANOBACTERIA AND CYANOTOXIN ECOLOGY IN LAKES AND DRINKING WATER by Chelsea Weirich The University of Wisconsin-Milwaukee, 2017 Under the Supervision of Professor Todd Miller, PhD Freshwater harmful algal blooms (FHABs) present a threat to ecological and public health in inland lakes. Problems associated with FHABs include human and animal illness, production of taste and odor compounds, and declining water quality and property values. Despite increased awareness and research on FHABs in recent decades, questions remain regarding long-term growth and diversity of FHABs in lakes that have taken measures toward slowing or reversing eutrophication, environmental factors impacting toxin occurrence, the most appropriate toxin analysis methods for protecting public health, and evidence for cyanotoxins detected in finished drinking water as a connection to exposure for potentially causing disease; this dissertation research seeks to add evidence to the field for answering such questions. Chapter 1 proposes maximum acceptable concentrations for a variety of cyanotoxins, in advance of recent US Environmental Protection Agency guidelines, and reviews childrens’ risk to cyanotoxin exposure, as well as possible clinical biomarkers. A long-term analysis of major FHAB-forming cyanobacteria and cyanobacterial community composition (CCC) in Chapter 2 demonstrates connections to organic nitrogen, changes in ice cover, total phosphorus, Schmidt stability and rainfall correlating with cyanobacterial abundance ii and CCC. Chapter 3 presents novel preservation methods for cyanotoxins in lake water samples for intercontinental shipping or storage, as well as describes patterns of occurrence for 13 cyanopeptides (e.g. microcystins, anabaenopeptins, cyanopeptolins, nodularin, microginin) in 22 lakes on a semi-global scale using liquid chromatography with tandem mass spectrometry quantification methods. These patterns in cyanotoxin occurrence are then analyzed based on characteristics of their lakes, including mussel invasion, dominant surrounding land usage, trophic status, and lake size. Finally, Chapter 4 characterizes amounts and types of cyanopeptides in raw and finished drinking water and efficiency of their removal among four drinking water treatment process trains. The resulting work demonstrates that other peptides are as common as microcystins in lakes and drinking water, and a variety of cyanopeptides at low doses are detectable in finished water from modern water treatment plants, while providing novel storage, extraction, and detection methods and defining parameters of interannual cyanobacterial dominance in a eutrophic lake with suggested future directions for monitoring. iii Dedication I dedicate these writings to my parents, Joseph Weirich and Catherine Breider, who encouraged me to study science with passion and persistence, and to my boyfriend, Nathaniel Kuhl, my source of endurance and inspiration. iv TABLE OF CONTENTS Dedication iv Table of Contents v List of Figures viii List of Tables ix List of Abbreviations xi Acknowledgements xv 1 Freshwater Harmful Algal Blooms: Toxins and Children’s Health 1.1 Introduction 1 1.2 FHAB toxins, mechanism of action, and symptoms 5 1.3 Children are at greatest risk for FHAB toxicosis 17 1.4 Exposure to FHABs through drinking water 24 1.5 FHAB toxins in food 30 1.6 Detecting algal toxins in clinical settings 31 1.7 Conclusions 35 2 Interannual Correlates of Cyanobacterial Community Composition and Total Biovolume over Two Decades in a Eutrophic Lake 2.1 Introduction 37 2.2 Study site description 41 2.3 Use of long-term datasets 42 2.4 Statistical analyses 46 2.5 Results 51 2.6 Factors Affecting Cyanobacterial Biovolume and Composition 61 v 2.7 Potential Effects of Climate Change 66 3 Large-Scale Spatiotemporal Perspective on Cyanotoxin Occurrence in Freshwater Lakes 3.1 Introduction 71 3.2 Sample collection 75 3.3 Reagents and Materials 76 3.4 Extraction of Cyanobacterial Peptides from Frozen Water Samples 79 3.5 Algal Toxin Detection Using Liquid Chromatographic Separation Coupled with Tandem Mass Spectrometry 80 3.6 Study Areas 81 3.7 Statistical Analyses 84 3.8 Spike-recovery analysis 85 3.9 Cyanopeptide occurrence 87 3.10 Cyanopeptides differentiated by environmental factors 92 3.11 Extraction Recovery 104 3.12 Cyanopeptide Preservation in Field Samples 105 3.13 Cyanopeptide Occurrence and Relations to Lake Characteristics 105 4 Cyanopeptides in Four Wisconsin Drinking Water Treatment Plants 4.1 Introduction 110 4.2 Drinking Water Source & Sample Collection 113 4.3 Cyanopeptide Extraction from Drinking Water Samples 120 4.6 Statistical Analyses 122 4.7 Cyanopeptide Detection Through Four DWTPs 123 vi 4.8 Daily Sampling at DWTP 4 129 4.9 Calculating Removal Efficiency of Cyanopeptides 131 4.10 Discussion 133 5 Conclusions and Future Directions for the Study of CyanoHABs and Cyanobacterial Secondary Metabolites 138 6 References 148 Appendix A: Chapter 2 Supplemental Data 190 Appendix B: Chapter 3 Supplemental Data 199 Appendix C: Reference Materials and Solutions 205 Appendix D: Detailed Protocols 210 Curriculum Vitae 237 vii LIST OF FIGURES Figure 1.1 Examples of structures of cyanobacterial toxins 11 Figure 2.1 Average biovolume of cyanobacterial taxa from 1995-2014 43 Figure 2.2 Dendrogram heat map of annual average BV for all cyanobacterial taxa counted in Lake Mendota 53 Figure 2.3 Detrended correspondence analysis correlating annual average cyanobacteria and environmental driver variables 55 Figure 2.4 Annual trends of environmental factors significantly correlated with cyanobacterial biovolume 58 Figure 3.1 Area, volume, trophic status, and max depth of lakes 83 Figure 3.2 Boxplot of cyanopeptides detected in all analyzed samples 88 Figure 3.3 Cyanopeptide detection in lakes in the United States 90 Figure 3.4 Cyanopeptide detection in lakes in Europe, Canada, Argentina 91 Figure 3.5 Average cyanopeptide concentrations by trophic state, land usage, and mussel colonization 96 Figure 3.6 Average cyanopeptide concentrations by elevation, lake mixing, and surface area 98 Figure 3.7 Average cyanopeptide concentrations by lake volume and depth 102 Figure 4.1 Lake Winnebago Pool 115 Figure 4.2 Treatment trains at Winnebago DWTPs 1 and 2 116 Figure 4.3 Treatment trains at Winnebago DWTPs 3 and 4 118 Figure 4.4 Mean cyanopeptides throughout drinking water treatment processes 134 Figure 5.1 Microcystins and Chlorophyll-a in Northern Lake Winnebago 140 viii LIST OF TABLES Table 1.1 Major FHAB toxins, target organs, symptoms, effect levels, and observed environmental concentrations 7-9 Table 1.2 Maximum acceptable concentration (MAC) in μg/L for common FHAB toxins 19 Table 2.1 Average Biovolume of Cyanobacterial Genera Detected More Than Five Years in Lake Mendota from 1995-2014 48 Table 2.2 Variables Used for Analysis of Long Term (1995-2014) Effects of Climate and Physical Limnology on Cyanobacterial Biovolume and CCC in Lake Mendota 49 Table 2.3 Variables Used for Analysis of Long Term (1995-2014) Effects of Nutrient Availability on Cyanobacterial Biovolume and CCC in Lake Mendota 50 Table 2.4 Significant Interannual Correlations Between Cyanobacterial Biovolume and Environmental Variables 57 Table 3.1 Characteristics of Samples Collected 77-78 Table 3.2 Descriptive Characteristics of Lakes Comparing Toxin Profiles 82 Table 3.3 Percent Recovery of Cyanobacterial Peptides from Tested Extraction Methods 86 Table 3.4 Mean, Standard Deviation, and Frequency of Cyanopeptide Detection 89 Table 3.5 Kruskal-Wallis Results Comparing Cyanopeptides Among Mesotrophic Lake Characteristics 94 Table 3.6 Cyanopeptides in Eutrophic and Hypereutrophic Lakes by Environmental Characteristics According to a Kruskal-Wallis Test 95 ix Table 4.1. Cyanopeptide Occurrence in Weekly Raw and Finished Water Samples from Four DWTPs 124 Table 4.2 Descriptive Statistics for Weekly Microcystin Occurrence After Each Plant Treatment Phase 125 Table 4.3 Descriptive Statistics for Weekly Cyanopeptolin Occurrence After Each Plant Treatment Phase 128 Table 4.4 Descriptive Statistics for Weekly Anabaenopeptin Occurrence After Each Plant Treatment Phase 180 Table 4.5 Descriptive Statistics for Cyanopeptides from All (Weekly and Daily) DWTP 4 Samples 130 Table 4.6 Percent Cyanopeptide Removal (PCR) After Major Treatment Phases 132 Table 5.1 Lillinonah Samples with Elevated Microcystins Not Associated with Poor Water Quality 142 x LIST OF ABBREVIATIONS AC – activated carbon ACE – angiotensin-converting enzyme ACN – acetonitrile